Compositions and methods for attracting mosquitoes and repelling sand flies

ABSTRACT

Compositions and methods for affecting dipteran hematophagous parasites. The compositions contain at least one dipteran semiochemical and at least one phagostimulant. The compositions may further include a pesticide. The semiochemical may be a floral attractant and the phagostimulants may be sugar-based. The compositions may be useful in attracting mosquitoes and/or repelling sand flies.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application No.62/078,039, filed on Nov. 11, 2014, and is a continuation in part ofU.S. patent application Ser. No. 14/938,696 filed on Nov. 11, 2015, theteachings of which are expressly incorporated by reference.

STATEMENT RE: FEDERALLY SPONSORED RESEARCH/DEVELOPMENT

Not Applicable

BACKGROUND

1. Field of the Invention

The present invention relates to products, systems, and methods forcontrolling adult populations of blood-feeding insects, both nuisancepests and vectors of disease. More specifically, this invention relatesto methods and systems for attracting multiple species of mosquitoes,for purposes of both population control and monitoring; and forrepelling New and Old World sand flies (vectors of leishmaniasis)through the use of a novel combination of floral plant volatilesemiochemicals with potent and varied behavioral effects. Thissemiochemical formulation can be employed in a broad range of means,including a monolithic lure and attractant-impregnated adhesive to bedeployed in monitoring traps, and a liquid formulation that can beblended with insecticide to create an attract and kill product amenableto spray application. It is also deployable in larger quantities instrategically placed, self-contained bait stations.

2. Background of the Invention

Insects of all species rely predominately on chemicals detected in theirenvironment for virtually every critical aspect of their lives, fromfemales' selection of appropriate sites upon which to deposit theireggs, location of desirable habitats and food sources and the avoidanceof undesirable ones, to the finding and selection of a mate. Thesebehavior-modifying chemicals, known collectively as semiochemicals, haveoften been used in attempts to manage or suppress insect pestpopulations through a wide variety of methods, such as mating disruption(artificial treatment of a vulnerable field or environment withsynthetic sex pheromone in such a way that the male insect is unable tolocate a mate within that field); the placement of an attractant in amonitoring trap or as part of a mass trapping program; repellency, todrive insects away from susceptible host organisms; and attract and kill(A&K), in which an attractant is applied in combination with a killingagent, typically a small quantity of insecticide, to draw insects to adefined location and kill them before they can either reproduce or causeany damage or disease to host organisms.

The pests targeted by the present invention, dipteran hematophagousparasites (those that feed on the blood of humans and animals and in sodoing transmit a broad range of blood-borne pathogens), include some ofthe most harmful insects to the health and prosperity of humanpopulations, both in the U.S. and abroad, that the world has ever known.Mosquitoes transmit some of the most devastating human diseases aroundthe world, such as malaria, arbovirus fevers such as dengue, yellowfever, chikungunya and West Nile virus, and filarial diseases, such aselephantiasis and river blindness among others. Vector-borne diseasestransmitted by insects are increasing in prevalence worldwide, and it islikely this trend will only increase in importance as climate patternschange in the future. According to the American Mosquito ControlAssociation, over one million people die from mosquito-borne diseaseseach year. West Nile virus is currently the mosquito-borne disease (MBD)of greatest concern in the U.S., with over 30,000 human cases reportedto the CDC since the disease was first detected in 1999. Historically,several other mosquito-borne diseases, including malaria, yellow fever,and dengue fever, were common killers in the United States. Persistentmosquito control efforts in this country and other industrializednations have protected their citizens from mosquito-borne illnesses fordecades. Although these efforts have been successful in keeping many ofus from contracting these illnesses, recent outbreaks of dengue fever,eastern equine encephalitis and the constant threat of West Nile virusare a sobering reminder of the constant necessity for effective,accurate mosquito control and monitoring techniques.

Sand flies also exert harmful impacts on people all over the globe,especially in under-developed regions, as vectors of leishmaniasis,caused by infection with Leishmania parasites. According to estimates bythe U.S. Centers for Disease Control and the World Health Organization,leishmaniasis is found in over 90 countries, putting over 310 millionpeople at risk. Approximately 300,000 cases of visceral leishmaniasis,the more severe form of the disease, are reported each year, resultingin over 20,000 deaths. While leishmaniasis is curable with propertreatment, most cases occur in impoverished regions, where access tohealth care is often limited. No vaccines are available to preventLeishmania infection. The best method of preventing the disease is toprevent contact between sand flies and their hosts, making effective,low-cost control technologies a major priority in affected areas.

In addition to the impacts biting dipterans have through their capacityto transmit disease, their blood-feeding behavior itself exerts harmfuleffects on productivity and quality of life for the people and animalsthey feed upon, particularly for outdoor workers, residents of rural orremote communities, and livestock species. Animals suffering fromattacks by large numbers of mosquitoes—under these circumstances,ruminants like cattle and sheep can lose as much as 300 mL of blood in asingle day—often do not feed properly, grouping together and attemptingto fend off mosquitoes rather than grazing, resulting in decreasedweight gains and reduced milk production in dairy cattle. There is ascarcity of recent data attempting to quantify these losses, but olderreports estimate economic losses of up to $61 million in a year as aresult of mosquito feeding on livestock.

Nuisance biting by hematophagous dipterans also negatively impacts humanproductivity and prosperity. Aside from the economic opportunity lost ifan individual contracts a vector-borne illness on the job, productivityamong outdoor workers has been shown to plummet when they are forced toshare their work environments with large numbers of mosquitoes. In somecases, agricultural employees may refuse to work where mosquitopopulations are high. Nuisance mosquitoes may also decrease propertyvalues; this is particularly evident in the southeastern U.S. Before theimplementation of organized mosquito control efforts in the state ofFlorida, coastal communities suffered such severe mosquito infestationsthat they were actually forced to close down during the summer months.Better mosquito control practices have not only fostered better qualityof life in these communities, but have also enabled further developmentof coastal regions, leading to an influx of tourism. There has beenfound to be a correlation between declining populations of Aedestaeniorhynchus, the saltmarsh mosquito, and increased touristexpenditures.

In light of these impacts, it is little surprise that there is anever-increasing demand for effective vector control products. The totalmarket size for mosquito adulticides (pesticide products specificallytargeting adult insects) alone is estimated at $4.8 billion. More than352 million acres are managed for vector control by approximately 1800mosquito control districts and municipalities in the U.S. at an averagecost of around $5/acre/day, a market size of approximately $4 billionper year. This is an expanding market, in the U.S. and globally, and iscurrently dominated by conventional chemical insecticides, particularlyby broad-spectrum pyrethroid and organophosphate sprays. Though thesetypes of products have produced good results in the past, delivering ahigh degree of mortality against multiple species of blood-feedingdipterans, they possess a number of disadvantages that make them anunsustainable pest management solution over the long term, bothenvironmentally and practically. First, as most conventionalinsecticides kill by contact, they must be blanket sprayed over allsurfaces where mosquitoes are thought to be present in order to provideeffective population control. This has led to a high degree of publicconcern regarding the negative effects these chemicals may have on theenvironment (persistence in the soil, contamination of water supply,spray drift, etc.) and on non-target species (i.e., people, beneficialinsects such as pollinators and natural enemies, livestock and companionanimals), especially when they are applied near human habitations.

Secondly, as relatively simple chemical toxicants with only a singlemode of action, all conventional insecticides developed thus far share acommon weakness: the potential for the population of the target insectto become less susceptible—and eventually, all but invulnerable—to themover time. This loss of sensitivity to a particular toxin, calledresistance, is more likely to occur when that toxin is applied over awide area, against multiple consecutive generations of the targetinsect, and begins when certain individuals within the population,having genes that make them more resilient to the chemical being used,survive exposure to it long enough to produce viable offspring. Theseoffspring then carry these genes on to the next generation, creating thebeginnings of a population that is resistant to the toxin.

Another critical component of an effective system of management forbiting dipterans is an accurate and timely method of populationmonitoring. Current monitoring traps tend to rely on a certain class ofsemiochemicals in order to attract mosquitoes and other blood-feedinginsects: host scent cues naturally produced by vertebrates, such ascarbon dioxide (CO2), which these insects use to orient toward apotential source for a blood meal. However, traps relying on CO2 as alure for dipterans can be extremely expensive ($300-$1400 for theinitial purchase), cumbersome, and challenging to maintain, requiring aconstant supply of electrical or battery power and frequent replenishingof the CO2 source in order to function effectively. They are alsoinherently limited in their capacity to attract the target insects: CO2and other attractants designed to mimic the scent profile of avertebrate host are only attractive to female dipterans seeking a bloodmeal. Scientists and vector control researchers have long focused onfemale blood feeding and methods of breaking transmission at this stage,but we believe this is an improper area of emphasis. Blood feedingbehavior, despite its importance to mosquitoes' impacts as diseasevectors, is actually a comparatively rare event in the insect's lifecycle. Female mosquitoes do not require a blood meal to nourishthemselves, only to complete development of their eggs, and so are onlyrequired to pursue this food source two or three times during theirlives, while males do not blood feed at all. A trapping program or anA&K strategy using CO2 or any other vertebrate host cue would thereforeonly have one or two opportunities to eliminate a mosquito before itbecomes capable of transmitting a pathogen (a minimum of two blood mealsis required for transmission to occur, one to acquire the pathogen froman infected individual, and one to introduce it to an uninfectedindividual).

As such, we have designed the current invention to exploit anotheraspect of hematophagous insect behavior, common to all mosquitoes, bothnuisance and vector species, regardless of gender, age, feedingpreference, or physiological status: their reliance on sugar solutionsto fuel their metabolism. Sugar-based food sources, mainly acquired fromflowers and extra-floral nectaries, are absolutely essential in order tosustain the highly active, highly mobile lifestyle that enablesmosquitoes and sand flies to survive and reproduce. As such, mosquitoesseek sugar meals on an almost daily basis throughout their lives,relying on floral-produced scent plumes to guide them to appropriatesources of nectar and plant juices, from the time they emerge as adultsto the time they die. A female mosquito will engage in thissugar-seeking behavior multiple times before ever pursuing a vertebratehost for blood feeding; she may take from eight to 12 sugar meals priorto her first blood meal, making her far more vulnerable to a floral orsugar-based attractant than to one intended to mimic the scent of apotential blood meal source. A floral plant volatile-based attractant,therefore, would have as many as 10 more opportunities to capture orkill a dipteran vector before it can transmit disease than an attractantmeant to manipulate blood-feeding behavior.

Our invention, called Vectrax, is designed as a highly potent andversatile blend of floral attractants and sugar-based phagostimulantsthat will act as an effective lure for all species of mosquitoes, ofeither gender, of all physiological states. This invention can beutilized in a wide variety of methods. Vectrax can be deployed alone tosubstantially improve monitoring efforts for vector mosquitoes of bothestablished and emerging MBDs in the U.S., or blended with smallquantities of insecticide to create an attract and kill (A&K)formulation.

For monitoring purposes, this attractant blend could be deployed as along-lasting monolithic lure, to be placed in virtually any type oftrap, or it could be blended directly into the adhesive for a stickytrap. Neither of these trap-lure systems would require a source of poweror a CO2 source in order to function, drastically reducing the effortand cost required to deploy and maintain the trap. This enhancedmonitoring capacity could help not only to improve strategic timing andlocation of mosquito control efforts (i.e., identification of keymosquito reproductive sites, targeting of high populations byinsecticide sprays), but also improve surveillance of mosquito-borneillnesses, both those already established, and those threatening toinvade from foreign regions. Current methods of surveillance of thesediseases are to a great extent reliant on case reports of both infectedpeople and animals—a relatively slow and ineffective indicator, at leastin terms of informing vector management decisions: mosquitoesresponsible for a given case of infection with a mosquito-borne pathogenare likely to be long gone from the area where transmission occurred bythe time the case is reported. While mosquitoes collected in CO2monitoring traps have also been used as a means to trackvector-transmitted illnesses, this method of surveillance can also beimpractical if the prevalence of the pathogen is low, such as would bethe case with a newly introduced pathogen. In such a situation, so fewmosquitoes within a given population would carry the disease agent thatit would require a very large sample size in order to detect itspresence. By increasing both the capacity to attract vector mosquitoes,and the number of monitoring traps that can be placed over the same areawithin the same time and budget constraints, Vectrax could substantiallyimprove the range and sensitivity of current monitoring programs forestablished and emerging mosquito-borne illnesses, both within the U.S.and abroad. In addition, while no trap design currently available hasdemonstrated the capacity to reduce mosquito populations or frequency ofbiting to any significant degree, the increased potency and decreasedcost of the Vectrax attractant compared to CO2 could enable a method ofpopulation control by mass trapping, by enabling deployment of traps ata high enough density for an effective mass trapping program.

Vectrax can also be blended with a small quantity of insecticide tocreate an A&K formulation. Broadly-defined, the A&K technique of pestcontrol consists of attracting adult males, females, or both sexes of apest species to an insect control agent (e.g., insecticide, sterilant,or insect pathogen). The insect attractant can be a chemical attractant,a visual cue, an acoustic cue, or a combination of these. A highlyeffective attractant and appropriate insecticide are indispensableingredients of an effective A&K product. For such a formulation to work,insect pests must be lured to a toxicant, which they must contact and/orfeed upon. Contact with the toxicant must then either kill the insector, at minimum, result in sublethal effects that preclude that insectfrom effectively performing behaviors that are essential to its survival(feeding behavior, escape responses, etc.), or the survival of itspopulation (effective courtship, mating success). The attractant must beat least as effective, if not more so, as attractants naturally presentin the environment, so that the A&K formulation successfullyout-competes them and lures the insect pest to the control agent. Inmany cases, the A&K also contains phagostimulants that induce the insectpest to consume the toxicant formulation. One way for A&K formulationsto outcompete existing, natural sources of the stimuli in the treatedenvironment, is by having point sources present at significantly higherdensities than the competing natural sources, and/or by beingsignificantly more attractive to the target pest.

Though both methods rely on chemical toxicants to suppress pestpopulations, A&K techniques present many advantages over cover sprays ofconventional insecticides. Attract and kill typically deploys smalleramounts of toxicants, often contained within discrete point sources andcoupled to a species specific attractant, reducing the likelihood ofnegative environmental and non-target effects. There are alsosubstantial economic benefits to the use of A&K over blanket pesticidesprays. Various attempts have been made to describe and quantify thenegative impacts that pesticides have on environmental and human health[58], accounting for the combined costs of all pesticides for eachcountry, and not only the costs of individual pesticides at a localscale. For example, Leach and Mumford (2008) developed a simple toolthat quickly assesses the indirect costs of individual pesticides basedon their particular toxicological and environmental behavior, providinga tool to rapidly estimate the environmental and public health impactsof pesticides in U.S. dollars/hectare/application. The model calculatesthe cost of a cover spray of Malathion 50% EC at $8.72/ha (this isexternal cost of an application, not the actual cost of the pesticide)while if the same pesticide were applied as an A&K bait, the costplummets to $0.04 per ha.

Despite these advantages, with few notable exceptions, the use offormulations baited with phytochemical attractants used by mosquitoesand other biting dipterans to locate sugar meals, remains largelyunexplored. Very little is known of the composition of the naturalvolatile plant semiochemical blends attractive to biting dipterans, andfew extracts or synthetic blends of these phytochemicals have beendeveloped for use in vector management formulations.

Furthermore, current A&K formulations lack rainfastness, and sun/UVprotection, and consequently have shorter field lives than desired.Their effectiveness is drastically reduced with the incidence of rain,and they invariably leak or drift, contaminating soils and waterways.Current A&K producers frequently create formulations that lack thespecies selectivity necessary to allow it to be applied in the fieldwithout causing environmental or ecological damage. While A&K may bevery selective (i.e., if insect sex pheromones are used as attractants),attractants with broad effects, such as plant kairomones, sugarsolutions, food fermentation residues, and their combinations, need tobe tested for their impact on non-target organisms.

The present invention addresses all these shortcomings in previous A&Kstrategies targeted toward biting dipterans. Through an extensive seriesof lab bioassays and semi-field mesocosm (large mosquito-proofgreenhouses) trials, we have successfully developed a number ofattractant blends that have proven so highly effective against threemajor vector genera of mosquitoes, Anopheles (vector of malaria, thedeadliest MBD of the modern world), Aedes (vector of Dengue fever, oneof the most prevalent MBDs in tropical regions), and Culex, (vector ofWNV and other arboviruses), that it out-competes natural plant odors andattractants. We have also developed an irresistible phagostimulant blendof sugars and proteins that causes the mosquito to feed continuously onthe formulation until it is fully engorged, even when the formulationcontains lethal doses of insecticide.

In addition to this high degree of efficacy as a mosquito attractant andphagostimulant, Vectrax also represents a substantial improvement overother forms of mosquito control in terms of safety, affordability, andlong-term sustainability. This formulation is composed entirely oforganic ingredients, for maximum safety to humans and the environment.As a thick, gel-like material applied in discrete point sources ratherthan a spray film that covers all surfaces, this A&K formulation isamenable to targeted, strategic application, allowing the user to selectapplication sites that may be expected to have the largest possibleimpact on the target (areas of high mosquito populations, key mosquitobreeding sites, etc.). This method of application reduces the overallquantity of insecticide required to be applied over a given area toachieve and maintain effective control (due to the powerful attractionand phagostimulation that the formulation exerts on the target pests),while eliminating the risk of spray drift and decreasing the likelihoodof contamination of the soil or water where it is applied. Vectraxpossesses a particularly valuable advantage over other forms ofpesticides in that it has demonstrated no negative impacts on thecritical pollinator species, Apis mellifera, the honey bee. Vectrax issurprisingly repellent to honey bees and other hymenoptera. Preliminarystudies showed that honey bees completely avoided the floral attractantwhen it was placed in their foraging zones: during 5 minute observationsof feeding stations containing 20% sugar solution, we observed anaverage of 33±5.8 bee visitations. There were zero visitations onfeeding stations containing the same 20% sugar solution spiked with aminiscule quantity (0.01%) of our floral attractant.

Vectrax as an A&K solution is a more economically sustainable pestmanagement solution than insecticide cover sprays, as well as a moreenvironmentally friendly one. Aside from requiring a lesser quantity ofinsecticide to be applied per unit area to achieve effective control,Vectrax can be formulated into a sprayable liquid formulation that willquickly solidify after application, to protect the active ingredientsfrom degradation and extend its field life. This results in a lowerfrequency of application to maintain mosquito population suppression,reducing the overall cost of the pest control strategy. Furthermore, asan attractant formulation amenable to tank mixing, designed to functionin combination with a wide variety of different insecticides, Vectrax isfar less susceptible to the development of resistance in the targetinsects: research has shown that rotation of different insecticides isan effective method of preventing the build-up of resistant pestpopulations, as the development of resistance to multiple classes oftoxins becomes increasingly unlikely. The tank-mixed Vectrax A&Ksolution can be applied in the field in one of two ways: 1) inrelatively large quantities contained within bait stations, which wouldbe particularly useful in areas of high pest population density; and 2)in a large number of much smaller point sources, as applied throughmanual or mechanical spraying equipment. This manner of applicationwould be of use in areas with lower mosquito population density, as thehigh density of point sources would dramatically increase the likelihoodthat the target insects will encounter, respond to, and consume theinsecticide-laced formulation.

While this invention was originally developed as an attractant forbiting diptera, the floral blend that lends Vectrax its efficacy provedto be extremely complex in chemical composition and behavioral effects,with different compounds eliciting different responses in insectsexposed to them. Certain components within this blend have demonstrateda strong repellent effect against New and Old World species of sandflies, opening up a new and unexpected avenue of pest control by meansof this invention. Through a series of dual-choice bioassays, a newcombination of plant volatile semiochemicals was identified, which actsas a powerful repellent against two vector species of sand flies,Lutzomyia longipalpis and Phlebotomus dubosqui (vectors ofleishmaniasis). This semiochemical blend proved more effective againstthis species than DEET (N, N-diethyl-meta-toluamide), currentlyconsidered the gold standard in insect repellents, even when the latterwas applied at a substantially higher application rate (1 mgsemiochemical blend vs. 1,200 mg DEET), suggesting that our inventionhas the capacity to repel sand flies more effectively with less materialand less cost required than topically applied products like DEET. Femalesand flies repelled from host environments will be less likely to obtaina blood meal, reducing egg production, while aggregation by males, whichtypically occurs near hosts, will also be impeded by the presence of arepellent. Both of these interventions will lead to a decrease inreproductive success within the treated area, and over time, a reductionin sand fly population size.

Regardless of the application method—whether as an attractant formosquitos or a repellent for sand flies, as an insecticide-free methodof pest control or blended with small amounts of reduced-risk toxicantsto create a more environmentally sound A&K formulation—this inventionwill diminish the impacts of biting diptera by a) reducing the frequencyof biting events, and consequently, opportunities for vector-bornedisease transmission to occur; b) reducing loss of productivity insusceptible livestock due to irritation and blood loss; and c) improveconditions at outdoor work sites, leading to greater worker comfort andproductivity. This is highly advantageous in any location wheremosquitoes create nuisance and health problems, but is particularlyvital in developing areas of the world, where methods of vector-bornepathogen prevention are still largely reliant on indoor interventions(i.e., insecticide-treated bed nets to prevent malaria transmission),leaving outdoor biting events a major source of transmission.

BRIEF SUMMARY

One embodiment of the present disclosure is directed toward acomposition for affecting dipteran hematophagous parasites. Thecomposition includes at least one dipteran semiochemical and at leastone phagostimulant. In particular, the semiochemical may be a floralattractant and the phagostimulant may be sugar-based. Further, thesemiochemical may be a mosquito attractant and/or a sand fly repellant.

The semiochemical and phagostimulant may be contained within asubstrate. In particular, examples of, and methods of making, suitablesubstrates are recited in U.S. Pat. No. 7,887,828 titled Dual ActionOrganic Formulation to Control Two Stages of Insect Pests, the entiretyof which is incorporated by reference herein. The substrate may, forexample, be a wax emulsion, microspheres, a latex solution, hot meltglue, a resin, or plastic flakes. In the case where the substrate is awax emulsion, it may be a wax carrier such as a paraffin wax, carnaubawax, beeswax, candelilla wax, fruit wax, lanolin, shellac wax, bayberrywax, sugar cane wax, microcrystalline wax, ozocerite, ceresin, montanwax, or combinations thereof. In a particular embodiment, the waxemulsion may include 30% by weight paraffin wax; 4% by weight soy oil;2% by weight sorbitan monostearate; 1% by weight vitamin E; and 58% byweight distilled water. In another embodiment, the wax emulsion mayinclude 45% by weight microcrystalline wax; 6% by weight soy oil; 3% byweight sorbitan monostearate; 1% by weight vitamin E; and 40% by weightdistilled water. In yet another embodiment, the substrate may be a hotmelt glue. The hot melt glue may be, for example, a polymer ofethylene-vinyl acetate, polyethylene, polypropylene, a polyamide, or apolyester.

In certain embodiments wherein it is intended to kill the dipteranhematophagous parasite, or another insect, the composition may furtherinclude a pesticide.

Examples of suitable compositions for use with the present disclosureare recited below in Tables 1-4:

TABLE 1 linalool 5%-25% by weight; phenylacetaldehyde 5%-45% by weight;β-myrcene 0%-50% by weight; citronella oil 5%-45% by weight; eucalyptol5%-25% by weight; geraniol 10%-20% by weight; camphene 5%-30% by weight;ocimene 1%-45% by weight; anethole 5%-45% by weight; anisic acid methylester 0%-25% by weight; phenethyl alcohol 1%-35% by weight;caryophyllene 2%-20% by weight; 4-methoxybenzyl alcohol 1%-15% byweight; methyl salicylate 0.1%-20% by weight; γ-terpinene 1%-45% byweight; α-terpinene 1%-45% by weight; limonene 5%-30% by weight; BHT1%-25% by weight; dipentene 1%-15% by weight; sugars 5%-60% by weight;thickeners 0.5%-5% by weight; preservatives 0%-2% by weight;antioxidants 0.1%-15% by weight; sunlight stabilizer 0.1%-10% by weight;wax 2%-40% by weight; emulsifier 0.5%-5% by weight; soybean oil 1%-40%by weight; and liquid carrier 1%-70% by weight.

TABLE 2 linalool 5%-25% by weight; phenylacetaldehyde 5%-45% by weight;β-myrcene 0%-50% by weight; citronella oil 5%-45% by weight; eucalyptol5%-25% by weight; geraniol 10%-20% by weight; camphene 5%-30% by weight;ocimene 1%-45% by weight; anethole 5%-45% by weight; anisic acid methylester 0%-25% by weight; phenethyl alcohol 1%-35% by weight;caryophyllene 2%-20% by weight; 4-methoxybenzyl alcohol 1%-15% byweight; methyl salicylate 0.1%-20% by weight; γ-terpinene 1%-45% byweight; α-terpinene 1%-45% by weight; limonene 5%-30% by weight; eugenolor clove oil 0.001%-50% by weight; guava oil or guava juice or guavaextract 0.001%-50% by weight; BHT 1%-25% by weight; dipentene 1%-15% byweight; sugars 5%-60% by weight; thickeners 0.5%-5% by weight;preservatives 0%-2% by weight; antioxidants 0.1%-15% by weight; sunlightstabilizer 0.1%-10% by weight; wax 2%-40% by weight; emulsifier 0.5%-5%by weight; soybean oil 1%-40% by weight; and liquid carrier 1%-70% byweight.

TABLE 3 linalool 5%-25% by weight; phenylacetaldehyde 5%-45% by weight;β-myrcene 0%-50% by weight; anethole 5%-45% by weight; anisic acidmethyl ester 0%-25% by weight; phenethyl alcohol 1%-35% by weight;caryophyllene 2%-20% by weight; 4-methoxybenzyl alcohol 1%-25% byweight; methyl salicylate 0.1%-20% by weight; γ-terpinene 1%-45% byweight; α-terpinene 1%-45% by weight; limonene 5%-30% by weight; and BHT1%-25% by weight.

TABLE 4 linalool 5%-25% by weight; phenylacetaldehyde 5%-45% by weight;β-myrcene 0%-50% by weight; anethole 5%-45% by weight; anisic acidmethyl ester 0%-25% by weight; phenethyl alcohol 1%-35% by weight;caryophyllene 2%-20% by weight; 4-methoxybenzyl alcohol 1%-25% byweight; methyl salicylate 0.1%-20% by weight; γ-terpinene 1%-45% byweight; α-terpinene 1%-45% by weight; eugenol or clove oil 0.001%-50% byweight; guava oil or guava juice or guava extract 0.001%-50% by weight;limonene 5%-30% by weight; and BHT 1%-25% by weight.

Another embodiment of the present disclosure is directed toward methodsof affecting dipteran hematophagous parasite populations. The methodincludes administering a composition to a region known or suspected tocontain dipteran hematophagous parasites, wherein the compositionincludes at least one dipteran semiochemical and at least onephagostimulant. The method may be used to attract mosquitoes. The methodmay be used to monitor the population of dipteran hematophagousparasites and/or to kill them by including a pesticide within or nearbyto the composition or by trapping the parasites at the site of thecomposition. The method may be used additionally, or alternatively, torepel sand flies. One benefit of the methods described herein is thatthe compositions do not negatively impact honey bee populations withinthe administered region. Another benefit of the described methods arethat the composition may be administered in numerous forms, includingbut not limited to, sprayable forms, monolithic lures, and sticky trapadhesives.

DETAILED DESCRIPTION

The detailed description set forth below is intended as a description ofthe presently preferred embodiment of the invention, and is not intendedto represent the only form in which the present invention may beconstructed or utilized. The description sets forth the functions andsequences of steps for constructing and operating the invention. It isto be understood, however, that the same or equivalent functions andsequences may be accomplished by different embodiments and that they arealso intended to be encompassed within the scope of the invention.

The compositions described in the present disclosure, coined Vectrax,are long-term slow release formulations that protect all incorporatedactive ingredients (the potent attractants, phagostimulants andinsecticides) from rain, decomposition and degradation. The dipteranpest control applications for which this formulation may be implementedare many and varied. By incorporating active ingredients (AIs) in such acontrolled release matrix, the effectiveness and longevity of these AIsin the field can be significantly improved, to such an extent that asingle application can provide weeks to months of protection against thetarget dipterans. As a result of this extended longevity and fieldhardiness, Vectrax can be deployed as a preventative measure, ahead ofoutbreaks of mosquitoes and other target biting dipterans. As aflowable, thick liquid formulation, this product is amenable tomechanical application through standard, readily available sprayequipment, but it effectively anchors to the substrate to which it isapplied, so that it doesn't dislodge during rain. Once the formulationhas solidified, it protects the AIs continuously, so that it sustainsconsistent and effective AI release over an extended period of time,ensuring full strength attraction, phagostimulation and/or insecticidaleffects, depending on the blend of components incorporated.

As floral- and sugar-based bait for mosquitoes, of both vector andnuisance species, so powerfully attractive to mosquitoes that itout-competes natural plant odors and attractants, Vectrax may be appliedalone, to substantially improve monitoring efforts for vector mosquitoesof both established and emerging mosquito-borne pathogens in the U.S.,by providing a lure that is as attractive or more so than currentCO²-baited traps at only a very small fraction of the cost. For example,one trial showed that within a single hour, a single passive trap,baited with a small quantity of variant of the Vectrax lure, capturedvirtually every mosquito released in test rooms (featureless 5×4×3 mtest rooms, temperature controlled at 27±3° C., humidity at 85±7%,lights off). An average of 98±1% (10 replicates) of 500 nulliparousAedes aegypti females were captured within this interval. This resultwas achieved with the use of a very simple, inexpensive, passive gluetrap design: a PVC tube (10.2 cm diameter, 30.5 cm tall) treated with apressure sensitive glue to capture mosquitoes upon first contact oftheir tarsi, and thus preserve captured specimens for furtheridentification and analysis. The enhanced monitoring capacity that couldbe achieved through the application of such a simple trap-lure systemcould help not only to improve strategic timing and location of mosquitocontrol efforts (i.e., identification of key mosquito reproductive sitesand targeting of high populations by insecticide sprays), but also toimprove surveillance of mosquito-borne illnesses, both those alreadyestablished in U.S. populations, such as WNV, and those threatening toinvade from foreign regions, like the recent detections of Chikungunyaand Dengue fever in the southern states.

Vectrax could also be blended with small quantities of insecticide tocreate an attract and kill (A&K) formulation that could be applied instrategic locations to draw mosquitoes away from potential hosts, bothhuman and animal, and kill them before they have a chance to bite. SinceVectrax is designed to function primarily with insecticides that work byingestion (requiring the target insects to actually consume theformulation in order to be effective), the formulation also containsextremely powerful sugar- and protein-based phagostimulants (feedingstimulants), which have been shown to induce target mosquitoes to feedcontinuously on Vectrax until fully engorged, even when the formulationcontains lethal doses of insecticide. In large semi-field trialsconducted in the large mesocosms at Ohio State University, a single 100mL point source of Vectrax, impregnated with 2% permethrin (by weight ofemulsion), applied to a single leaf within the mesocosm, successfullyattracted and killed roughly half of the 500 virgin two-day-old femaleAnopheles gambiae mosquitoes released the first night after theirrelease, despite the presence of many other plants known to beattractive to sugar-seeking mosquitoes. The remaining half of thereleased mosquitoes were killed during the second night, eliminatingvirtually every mosquito in the treated mesocosms within 48 hours. Incontrast, more than 90% of the mosquitoes released in the controlmesocosms remained alive at the end of the second day following theirrelease.

In addition to this exceptional level of efficacy, careful design of theattractant and phagostimulant AIs, as well the various components of thecontrolled release emulsion, has resulted in a product that can be tankmixed with a broad range of registered pesticides, ensuring that Vectraxwill be adoptable in virtually any location, regardless of whattoxicants are most effective against a given species in a givenenvironment or situation. This adaptability also presents a considerableadvantage in terms of long-term applicability: because Vectrax canfunction effectively as an A&K system with so many different types ofinsecticides, it is nearly invulnerable to the development of resistancein the target insects. It is also a more sustainable method for thecontrol of dipteran pests than traditional insecticide cover sprays.Vectrax is composed entirely of organic ingredients, for maximum safetyto humans and the environment, even when toxicants are incorporated. Byselecting only reduced-risk insecticides, those that work as stomachpoisons rather than simply by contact, chances are lessened thatnon-target species will be harmed: only insects that are attracted tothe Vectrax point source and actually feed upon the material will suffernegative effects. Of particular importance and value is Vectrax's lackof negative effects on honey bee (Apis mellifera) populations,especially given the severe and still largely unexplained decline inhoney bee populations around the world. Vectrax is surprisinglyrepellent to honey bees and other Hymenoptera. Preliminary studiesshowed that honey bees completely avoided the floral attractant when itwas placed in their foraging zones: during 5-minute observations offeeding stations containing 20% sugar solution, an average of 33±5.8 beevisitations was observed. There were no visitations to feeding stationscontaining the same 20% sugar solution spiked with 0.01% of our floralattractant. Finally, Vectrax will be designed to reduce the exposure ofthe pesticide in the environment by retaining, protecting and slowlyreleasing the AI from discreet Vectrax point sources (instead of theblanket cover sprays used for conventional insecticides). These pointsources become beacons for the target species, and are easily avoidableby non-targets. Alternatively, larger quantities of Vectrax can bedeposited in bait stations, which would be particularly useful in areasof high pest population density.

Vectrax can also be deployed as a vehicle for the controlled release ofrepellent plant volatiles, to manage sand fly populations. Althoughoriginally evaluated as a sand fly attractant, as it was for mosquitoes,ISCA Technology scientists discovered a number of volatile plant-derivedcompounds that have demonstrated an extremely high degree of repellencyagainst both New and Old World sand flies, using only miniscule AIquantities, and in lab tests it has been observed to exert a “haloeffect” of repellency not seen in topically applied products. In aseries of dual-choice bioassays, in which two leishmaniasis vectors,Lutzomyia longipalpis and Phlebotomus dubosqui, were given a choicebetween a chamber treated with a variant of Vectrax and a controlchamber, both species overwhelmingly chose the control (95% L.longipalpis; 100% P. dubosqui), demonstrating strong repellency. Incomparison, the same experiment conducted with N,N-diethyl-meta-toluamide (DEET), considered the gold standard in insectrepellents, demonstrated weaker repellency to P. dubosqui thanVectrax—only 79% flies selected the untreated over the treated chamber,despite a higher application rate (1,200 mg DEET vs. 1 mg Vectrax). Thisplant-based repellent formulation could reduce sand fly populations in anumber of ways. Female sand flies repelled from host environments byVectrax will be less likely to obtain a blood meal, reducing eggproduction, while lek formation by males, which typically occurs nearhosts, will also be impeded by the presence of a repellent. Both ofthese interventions will lead to a decrease in reproductive successwithin the treated area, and over time, a reduction in sand flypopulation size. The volatile nature of this blend suggests that whenincorporated into a controlled-release formulation, it will act overgreater distances than current repellent formulations (e.g. DEET)—and istherefore capable of protecting an entire area from sand flies, ratherthan a single individual—and will remain active for longer periods.

Vectrax formulations belong to a “matrix-type” or “monolithic” categoryof controlled-release devices. These monolithic dispensers are definedas devices where the active ingredient (AI) is dispersed or dissolved ina polymer matrix. Release of the AI from a monolithic device occurs bydiffusion and can be described macroscopically by Fick's Law, whichstates that the movement of a molecule by diffusion is directlyproportional to the concentration of that molecule in a system.Microscopically, if one follows the movement of a molecule of an activeagent through a matrix, this molecule begins its journey in one of twoways. If it is dispersed in the matrix, it begins its journey bydissociating from other molecules in its crystal cell and solubilizinginto the polymer phase. If it is dissolved in the matrix, then this stepis bypassed. The molecule then diffuses through amorphous regions in thematrix that comprise the free volume of the system. The molecule canmove through the matrix in one of two ways. If it is very small comparedto the size of the amorphous spaces in the matrix, then it will diffusethrough the matrix by moving from one such space to another. If it isvery large compared to the size of those spaces, then segments of thepolymer comprising the matrix will have to be rearranged for diffusionof the active agent molecule to occur. Crystalline regions in the matrixare virtually impermeable to molecules of the active agent. Uponreaching the surface of the matrix, it will be released into theenvironment. A series of factors influence the rate of release of anactive agent from a monolithic device and include properties of thematrix material as well as properties of the active agent. Thetemperature of the matrix influences release of the active agent; athigher temperatures the free volume is increased, and diffusion occursfaster. At lower temperatures, the free volume is decreased, anddiffusion is slower. The thermal history of a polymer can also increaseor decrease the free volume of the system and lead to changes in thediffusional rate of an active agent. The property of the AI having thegreatest influence on its release rate is its molecular weight.Generally, larger molecules take more time to make their way through thefree space of a matrix. The partition coefficient of the active agentbetween the matrix and the environment can also influence the releaserate of that agent. If the agent readily partitions to the environment,then its rate of release will be diffusion-controlled and first order.If, however, partitioning of the active agent to the environment isrelatively slow, then its partition coefficient will determine itsrelease rate from the matrix, and the device will exhibit zero-orderrelease kinetics. The partitioning of the AI to the environment is afunction of its solubility in the matrix; compounds more soluble in thematrix partition to the environment more slowly. Vectrax emulsions in afield environment exhibit diffusion-controlled release. The surface areaof the device also influences its release rate. Vectrax dispensers withlarger surface areas release AIs at faster rates. The release rate of aVectrax formulation containing a fixed amount of semiochemical can bemodulated simply by changing a few parameters of the formulation, whichinclude the type of components used (e.g., wax composition,emulsifiers), their proportion in the formulation (e.g., percentage ofwater, oil or wax), the stage in manufacturing when different componentsare added, rheology, and finally, the characteristics of the dispenserupon application in the field (e.g., applied as microdollops of 1-10 μgeach or larger dollops of 1-5 g each).

Application Methods for the Invention:

Vectrax for Mosquitoes.

Vectrax can be deployed to manage nuisance and vector mosquitopopulations in outdoor environments in three ways: 1) for monitoringpurposes, when applied as a lure in virtually any form of trap; 2) inself-contained bait stations, either as an attractant alone, to draw theinsects away from important or vulnerable areas, or in combination withan insecticide to attract and kill them, permanently removing them fromthe environment; and 3) as a sprayable A&K formulation that can beapplied manually or through a wide variety of mechanized equipment,directly to foliage within the mosquitoes' habitat. Though thisformulation highly flexible in terms of application rate and method, aguideline application procedure for each method described above isincluded below.

Monitoring. Vectrax (attractant only) may be applied as a lure either byitself, or as a complement to any other type of attractant, includingCO₂, in virtually any type of trap. To use the formulation in this way,a small quantity of Vectrax, shaken or stirred to ensure that allincorporated AIs are in suspension, is deposited onto a stablesubstrate, such as a cotton ball or a segment of cotton gauze, and thensecured within the trap, typically through the application of some formof adhesive. The applied point source may range in size from a few μg to10s of grams in size, depending on a) the concentration/dilution of theattractant, and b) how long the attractant is desired to maintain itsactivity in the field. Alternatively, a quantity of the Vectraxattractant could be blended directly into the adhesive itself, beforebeing applied within the trap. Blending may be accomplished through awide variety of manual or mechanized mixing equipment. Vectrax-baitedtraps may be deployed at single locations, in order to lure mosquitoesaway from sensitive areas (i.e., rural residences, backyards,recreational activity sites) and remove them from the environment bytrapping them, or as part of a mosquito management strategy, placed atwhatever locations and whatever density is thought to be required toeffectively suppress mosquito populations.

Vectrax Bait Stations. To apply Vectrax within bait stations, theformulation—again, previously shaken or stirred—is loaded into areservoir that provides the target insects with easy access to theattractant material, similar to a hummingbird feeder. The attractantformulation may be applied by itself or blended with a small quantity ofan appropriate insecticide (the insecticide cyazypyr has proven quiteeffective in laboratory and mesocosm trials). In order to maintainmaximum A&K efficiency, the toxicant component should be blended intothe Vectrax attractant formulation as close to the time of applicationas possible. The attractant or A&K formulation may be loaded into thebait station in amounts ranging from a few grams to several liters,depending on the desired field life. If desired, a sponge or otherabsorbent material may be applied with the formulation, to facilitatelanding by the mosquitoes. As with Vectrax-baited traps, Vectrax baitstations may be deployed singly, as a means to divert or attract andkill mosquitoes in sensitive environments (i.e., rural residences,backyards, recreational activity sites), or as part of a mosquitomanagement strategy, placed at whatever locations and whatever densityis thought to be required.

Sprayable A&K. As with a Vectrax A&K formulation deployed in baitstations, a formulation intended to be applied directly to foliage as aliquid material should be blended with its toxicant componentimmediately prior to its application. Once the formulation is thoroughlyblended, it may be applied by hand, using simple tools such as knives,spatulas, brushes, or syringes; or mechanically, using anything from abackpack sprayer to a tractor or aerial spray equipment. The rate andquantity at which the formulation should be applied will depend on theneeds of the specific pest management situation, such as the desiredfield longevity (larger point sources will maintain their activity for alonger period than smaller point sources) and the density of the pestpopulation at the site, but for many studies examining Vectrax as an A&Kproduct, an application rate of 1 liter per hectare has proven adequate.

Vectrax for Sand Flies.

To apply Vectrax as a spatial and contact repellent for New or Old Worldsandflies, the formulation may be applied in an identical manner as forbait stations for mosquitoes or sprayable liquid formulation (see above)though in this case, these measures will serve the opposite purpose ofrepelling the target pest away from treated sites, rather than drawingthe insects to them for removal or monitoring. Because of this dualactivity, in areas treated with the Vectrax sand fly repellent wheremosquitoes are also present, a small amount of insecticide activeagainst biting Diptera can be blended with the formulation prior toapplication. Once the formulation has been properly blended, it may beapplied near sensitive sites, such as the outdoor area surrounding ahome, outdoor worksites known to be heavily infested with sand flies, orkey sand fly reproductive sites. When deploying the repellent in aself-contained repellent station, the amount of formulation to beapplied may range from a few grams to several liters, as with mosquitobait stations, depending on the desired field longevity. When applyingas a spray over larger areas, application rates/quantities are similarlyflexible, but an application rate of 1 kg per acre has proven effectivein most previous tests.

Experimental Results Containing Insecticide:

Materials:

Two formulations 250 mls each were prepared: i) Control formulationcontaining blank vectrax, and ii) Treatment formulation containingvectrax+insecticide (permethrin). Cotton wool. Thatching materials(bamboo leaves). Plastic sheet.

Procedure:

One hour before the release of mosquitoes, i) a plastic sheet was laidon the roof of each respective control and treatment Hut, ii) Thatchingmaterials were fixed on top of the plastic sheet for both huts incontrol and treatment spheres (the plastic sheet was to preventcontaminations of original thatching materials of the huts), and iii)Then cotton wool soaked in the blank vectrax for control sphere andvectrax+insecticide for the treatment sphere were placed on paper cupsturned upside down to six stations of this kind in both spheres

Mosquitoes Release:

At 10 AM on the day of release, 1 to 2 days old female An. gambiae sswere separated at equal numbers in the cages and starved until the timeof release (1 to 2 days old because at that age sugar is the preferredmeal). At eight hours of starvation, the mosquitoes were released inboth spheres at an equal number. The release was made by placing thecage at one corner of the sphere and providing a small exit from thecage (approximately 15 cm). Then mosquitoes were left in hut for 24 hrsso that they could acclimatize with the Vectrax control or TreatmentVectrax. After 24 hrs, a human subject went in the respective nets inhuts and counted the number of mosquitoes coming to bite. Therecapturing went on for three days following the release. Scores werethen recorded and totaled.

The above description is given by way of example, and not limitation.Given the above disclosure, one skilled in the art could devisevariations that are within the scope and spirit of the inventiondisclosed herein, including the use of various semiochemicals andpesticides to achieve the same intended effect. Further, the variousfeatures of the embodiments disclosed herein can be used alone, or invarying combinations with each other and are not intended to be limitedto the specific combination described herein. Thus, the scope of theclaims is not to be limited by the illustrated embodiments.

What is claimed is:
 1. A composition for attracting mosquitoes andrepelling sand flies, the composition comprising: linalool 5%-25% byweight; phenylacetaldehyde 5%-45% by weight; β-myrcene 0%-50% by weight;anethole 5%-45% by weight; anisic acid methyl ester 0%-25% by weight;phenethyl alcohol 1%-35% by weight; caryophyllene 2%-20% by weight;4-methoxybenzyl alcohol 1%-25% by weight; methyl salicylate 0.1%-20% byweight; γ-terpinene 1%-45% by weight; α-terpinene 1%-45% by weight;limonene 5%-30% by weight; and BHT 1%-25% by weight.


2. The composition of claim 1 further comprising: eugenol or clove oil0.001%-50% by weight; and guava oil or guava juice or guava extract0.001%-50% by weight.


3. A composition for attracting mosquitoes and repelling sand flies, thecomposition comprising: linalool 5%-25% by weight; phenylacetaldehyde5%-45% by weight; β-myrcene 0%-50% by weight; citronella oil 5%-45% byweight; eucalyptol 5%-25% by weight; geraniol 10%-20% by weight;camphene 5%-30% by weight; ocimene 1%-45% by weight; anethole 5%-45% byweight; anisic acid methyl ester 0%-25% by weight; phenethyl alcohol1%-35% by weight; caryophyllene 2%-20% by weight; 4-methoxybenzylalcohol 1%-15% by weight; methyl salicylate 0.1%-20% by weight;γ-terpinene 1%-45% by weight; α-terpinene 1%-45% by weight; limonene5%-30% by weight; BHT 1%-25% by weight; dipentene 1%-15% by weight;sugars 5%-60% by weight; thickeners 0.5%-5% by weight; preservatives0%-2% by weight; antioxidants 0.1%-15% by weight; sunlight stabilizer0.1%-10% by weight; wax 2%-40% by weight; emulsifier 0.5%-5% by weight;soybean oil 1%-40% by weight; and liquid carrier 1%-70% by weight.


4. The composition of claim 3 further comprising: eugenol or clove oil0.001%-50% by weight; and guava oil or guava juice or guava extract0.001%-50% by weight.